Chemical fingerprinting of wood sampled along a pith-to-bark gradient for individual comparison and provenance identification

Background and Objectives: The origin of traded timber is one of the main questions in the enforcement of regulations to combat the illegal timber trade. Substantial efforts are still needed to develop techniques that can determine the exact geographical provenance of timber and this is vital to counteract the destructive effects of illegal logging, ranging from economical loss to habitat destruction. The potential of chemical fingerprints from pith-to-bark growth rings for individual comparison and geographical provenance determination is explored. Materials and Methods: A wood sliver was sampled per growth ring from four stem disks from four individuals of Pericopsis elata (Democratic Republic of the Congo) and from 14 stem disks from 14 individuals of Terminalia superba (Cote d'Ivoire and Democratic Republic of the Congo). Chemical fingerprints were obtained by analyzing these wood slivers with Direct Analysis in Real Time Time-Of-Flight Mass Spectrometry (DART TOFMS). Results: Individual distinction for both species was achieved but the accuracy was dependent on the dataset size and number of individuals included. As this is still experimental, we can only speak of individual comparison and not individual distinction at this point. The prediction accuracy for the country of origin increases with increasing sample number and a random sample can be placed in the correct country. When a complete disk is removed from the training dataset, its rings (samples) are correctly attributed to the country with an accuracy ranging from 43% to 100%. Relative abundances of ions appear to contribute more to differentiation compared to frequency differences. Conclusions: DART TOFMS shows potential for geographical provenancing but is still experimental for individual distinction; more research is needed to make this an established method. Sampling campaigns should focus on sampling tree cores from pith-to-bark, paving the way towards a chemical fingerprint database for species provenance

Comparison of species classification models of mass spectrometry data : kernel discriminant analysis vs. random forest : a case study of Afrormosia (Pericopsis elata (Harms) Meeuwen)

Rationale: The genus Pericopsis includes four tree species of which only Pericopsis elata (Harms) Meeuwen is of commercial interest. Enforcement officers might have difficulties discerning this CITES-listed species from some other tropical African timber species. Therefore, we tested several methods to separate and identify these species rapidly in order to enable customs officials to uncover illegal trade. In this study, two classification methods using Direct Analysis in Real Time (DART) ionization coupled with Time-of-Flight Mass Spectrometry (DART-TOFMS) data to discern between several species are presented. Methods: Metabolome profiles were collected using DART ionization coupled with TOFMS analysis of heartwood specimens of all four Pericopsis species and Haplormosia monophylla (Harms) Harms, Dalbergia melanoxylon Guill. & Perr. Harms, and Milicia excelsa (Welw.) C.C. Berg. In total, 95 specimens were analysed and the spectra evaluated. Kernel Discriminant Analysis (KDA) and Random Forest classification were used to discern the species. Results: DART-TOFMS spectra obtained from wood slivers and post-processing analysis using KDA and Random Forest classification separated Pericopsis elata from the other Pericopsis taxa and its lookalike timbers Haplormosia monophylla, Milicia excelsa, and Dalbergia melanoxylon. Only 50 ions were needed to achieve the highest accuracy. Conclusions: DART-TOFMS spectra of the taxa were reproducible and the results of the chemometric analysis provided comparable accuracy. Haplormosia monophylla was visually distinguished based on the heatmap and was excluded from further analysis. Both classification methods, KDA and Random Forest, were capable of distinguishing Pericopsis elata from the other Pericopsis taxa, Milicia excelsa, and Dalbergia melanoxylon, timbers that are commonly traded

Pith-to-bark profiles of xylem vessel traits reveal unique information on tree performance in a tropical moist semi-deciduous forest of the Congo Basin

Xylem vessel features are the most investigated anatomical traits of Angiosperm wood. They are being analyzed in ecophysiological projects aiming at understanding sap flow and vulnerability to drought stress. Since long, size, grouping and density of vessels are also used as one of the first steps in timber identification, as they can easily be observed even with a hand lens. Vessels are also known to be related to tree height. The volumetric sap flow rate is inversely related to the length of the path, so that vessels need to widen at lower parts of the tree and taper towards the higher parts to assure optimal transport and a stable hydraulic resistance with progressing growth in height. Vessels are expected to narrow from roots to stem and further to the branches and the petioles. Accordingly it is also predictable that vessel features on pith-to-bark profiles are a proxy for height growth for individual trees. Since the functional groups of species of a tropical rainforest are based on height grow rate (related to light needs), it is expected that pith-to-bark profiles of vessels reveal information on temperament of the species. A methodology has been developed to establish pith-to-bark profiles of vessel features, based on long microtomic sections, image analysis and machine learning. Entandrophragma was used as a model genus. The differences between the four most common species of this genus have been explored. Wilcoxon signed rank test indicate that the difference in mean vessel size is significant between every species combination except between Entandrophragma candollei and Entandrophragma cylindricum. Interesting to note is the large vessel size range possible for Entandrophragma utile. Currently, pith-to-bark vessel trends are constructed that could help explaining difference in growth strategy. The approach offers appealing perspectives to find a formal way for sub setting tree species into functional groups and develop indexes for growing conditions of forest sites

When xylarium and herbarium meet : linking Tervuren xylarium wood samples with their herbarium specimens at Meise Botanic Garden

Background: The current data paper aims to interlink the African plant collections of the Meise Botanic Garden Herbarium (BR) and the Royal Museum for Central Africa Xylarium (Tw). Complementing both collections strengthens the reference value of each institutional collection, as more complete metadata are made available and it enables increased quality control for the identification of wood specimens. Furthermore, the renewed connection enables the linking of available wood trait data with data on phenology, leaf morphology or even molecular information for many tree species, allowing assessments of performance of individual trees. In addition to studies at the interspecific level, comparisons at the intraspecific level become possible, which could lead to important new insights into resilience to and impact of global change, as well as biodiversity conservation or forest management of Central African forest ecosystems. New information: By interlinking the Tervuren Xylarium Wood database with the recently digitised herbarium of Meise Botanic Garden, we were able to establish a link between 6,621 xylarium and 9,641 herbarium records for 6,953 plant specimens. Both institutional databases were complemented with reliable specimen metadata. The Tervuren xylarium now profits from taxonomic revisions made by botanists at Meise Botanic Garden and a list of phenotypical features for woody African species can be extended with wood anatomical descriptors. New metadata from the Tw xylarium records were used to add the country of collection to 50 linked BR herbarium specimens for which this information was missing. Furthermore, metadata available from the labels on digitised BR herbarium specimens was used to update Tw xylarium records with the date of collection (817 records), collection locality (698 records), coordinates (482 records) and altitude (817 records). In conclusion, we created a reference database with reliable botanic identities which can be used in a range of studies, such as modelling analyses, community assessments or trait analyses, all framed in a spatiotemporal context. Furthermore, the linked collections hold historical reference data and specimens that can be studied in the context of global changes

Assessing the natural durability of xylarium specimens : mini-block testing and chemical fingerprinting for small-sized samples

The resistance of wood against fungal decay, in short the natural durability, is one of the main criteria for defining the potential use of a wood species. Wood collections, or xylaria, offer the unique opportunity to screen many specimens and species for the latter purpose yet sample size is often limited and standardized tests are often not possible neither desired given the historical and cultural value of these specimens. Two different methods to determine the natural durability are tested and presented here, more specifically the mini-block test and chemical fingerprinting by Direct Analysis in Real-Time Time-Of-Flight Mass Spectrometry (DART TOFMS). Fungal decay by Trametes versicolor was determined for 577 mini-blocks collected from xylarium specimens and 602 mini-blocks from commercial species, not belonging to the xylarium collection, were included as a benchmark. Mass loss percentages of the different species are similar to reported values, supporting the use of the mini-block test when standardized testing is hardly feasible. Furthermore, as expected there is also a significantly negative relationship between density and the mass loss percentages from the mini-block test (r-Spearman = - 0.65***). Finally, partial least square-based prediction of recorded mass loss by using the DART TOFMS chemical fingerprints is promising (R-2-adjusted = 0.40***), yet the accuracy differs between species

Methodology for the treatment of increment cores to assess the vessel size distribution of a tree along a pith-to-bark transect

The sampling of increment cores constitutes a non-destructive approach to study the wood anatomy of a tree along a radial transect. The distribution and variability of anatomical features along such a transect can serve as a proxy of a tree’s growth dynamics. It is therefore valuable to develop methods of accessing and analyzing these data, such as the distribution of vessel sizes along a pith-to-bark transect. The processing of the increment cores begins with the production of long cross sections, 10 to 20 µm thick, using a microtome. The cross sections are then stained to highlight the anatomical features. Next, the cross sections are digitized using the Stream Motion software (Olympus Optical Co. Japan) on a scanning stage (SCAN 100 x 100, Märzhäuser Wetzlar, Germany) to create high resolution images. A pre-treatment of the images, with an image manipulation program such as GIMP 2 (www.gimp.org), is then required to enable their use for data analyses. This includes associating fragments to reconstruct a linear and continuous cross section, since it is difficult to consistently prevent the formation of tearing when cutting the cores. The data contained in the cross section images is finally accessed using the ImageJ software (www.imagej.net). Data such as the position, size, and vessels density along a pith-to-bark transect are then acquired and can therefore be used to study the physiology and growth dynamics of the trees. ImageJ features such as the Trainable Weka Segmentation plugin offer the opportunity to automatize the analyses, therefore allowing for a larger sampling size and faster processing

A new method for exploring and explaining dimensional stability of commercial and lesser-used wood species from the Congo Basin

One of the main criteria for assessing the potential use of a timber species is the dimensional stability or resistance against swelling/shrinkage. Dimensional changes due to change in moisture content can lead to several defects, e.g. warping, checking, splitting… Thus to support a fit for purpose mentality we need to quantify the dimensional stability of a species. We therefore propose a methodology based on Digital Image Correlation (DIC) to determine the dimensional stability and we will verify whether this methodoloy works for both standardised and non-standardised samples. Specimens were collected from the Tervuren xylarium (Royal Museum for Central Africa, Belgium), which contains over 80000 specimens comprising more than 13000 species (mainly Central Africa), a result of more than one century of collecting efforts, and can be considered as a sleeping beauty in terms of wood technological output. The xylarium offers opportunities to screen a large number of species if a DIC method could be developed, adapted to the various formats of xylarium samples. The axial plane of the specimens was sanded and a 0.5 cm thick section was sawn off to retrieve a sample for DIC. The methodology is based on the calculation of pixel displacements using flatbed scans (SilverFast SE Plus 8 – scanner at 2400 dpi and stored as 48 bit colour images) from the same sample (cross section transversal plane) taken at different humidities (60 % RH – 90 % RH). The results from the DIC method are compared with conventional caliper (Absolute digimatic caliper, Mitutoyo, 0.01 mm) measurements on standardised samples (5.0 x 5.0 x 0.5 cm, 89 samples, 10 species). Wood density (kg/m³) (ovendry/volume) was determined as well on the same specimens using the Archimedes principle. In total 421 non-standardised samples (comprising 53 species) were analysed. The results of the calliper measurements and the DIC method are very similar (linear model: R² adjusted - tangential = 0.96, R² adjusted - radial = 0.94) showing the potential of the digital image correlation method. This method also shows potential towards determining whether the deformation is uniform or rather local. The correlation between average volumetric swelling and average wood density per species was rather weak but significant (r-Pearson = 0.36, p ≤ 0.01). Afzelia species, Pterocarpus tinctorius and Millettia stuhlmannii have a high density and high dimensional stability, thus interesting for exterior joinery applications. On the other side of the spectrum, Antiaris toxicaria and Canarium schweinfurthii have a very low wood density and high volumetric swelling, showing less potential to be used as a high quality timber species. The successful results of this DIC methodology allow for a large screening of non-standardised specimens present in xylaria worldwide